A peach ethylene response factor PpERF61 is involved in fruit ripening by modulating ripening-related genes and PpSEP1

Fruit ripening is a highly coordinated process at the transcriptional level, requiring a hierarchical regulatory network formed by transcription factor and ripening-related genes. MADS-box transcription factors are core regulators that play a key function in the ripening process of fruit, targeting a large amount of downstream ripening-related genes. However, the upstream regulators of MADS-box transcription factors are poorly understood. In this study, we identified an ethylene response factor PpERF61 by yeast one-hybrid screening with the promoter of PpSEP1 (MADS-RIN homologs). The expression pattern of PpERF61 was closely associated with ethylene biosynthesis and fruit softening across six peach cultivars. The transient overexpression and VIGS assay showed that overexpression or silencing of PpERF61 in peach promoted or inhibited fruit softening, while ethylene production was increased or decreased, respectively, demonstrating that PpERF61 has effect on ethylene generation and fruit softening during peach ripening. As a nuclear-localized transcription factor, PpERF61 activated the ripening-related genes transcription, including PpACO1, PpACS1, PpPL1, and PpPL15, by binding directly to their promoters. Interestingly, PpERF61 acts as upstream of PpSEP1 and binds directly to the PpSEP1 promoter, thereby forming PpERF61-PpSEP1 module to promote the PpPG2 and PpPG3 transcription. Based on these findings, we conclude that PpERF61 positively regulates ethylene biosynthesis and fruit softening by directly activating ripening-related genes or activating the PpERF61-PpSEP1 signaling cascade. Collectively, these results enrich knowledge of the complex cascade network for peach fruit ripening.